Serotonin N-Acetyl-Transferase Inhibitors for Circadian Rhythm Disorders - Circadian rhythm (CR) dysregulation contributes to mental health disorders, including major depressive disorder (MDD), bipolar disorder (BD), and seasonal affective disorder (SAD). Melatonin has been strongly associated with CR, but despite years of research, many questions remain regarding its role and how it influences mood. The rate-limiting step in melatonin synthesis involves the enzyme serotonin-N-acetyltransferase (SNAT, AANAT). Inhibition of SNAT would be a valuable approach for studying the physiological function of melatonin and could be used to treat disorders such as SAD that involve abnormally high melatonin. Published inhibitors have problems with cell permeability, selectivity, and/or potency, which have prevented advancement to testing in humans. The aims of this project are 1) to identify potent, selective and cell permeable SNAT inhibitors by virtual screening, and 2) to use structure-guided design to improve potency of cell permeable SNAT inhibitors. To achieve Aim 1, we will use a more physiologically relevant SNAT structure than has been used in the past, bound to its chaperone, with a more closed binding site, which we hypothesize will have greater predictability than previous models for screening commercial compounds in the ZINC database. As a selectivity filter, we will dock commercial compounds with high scores for predicted SNAT binding against the anti-targets, melatonin receptor 1 and 2 (MT1-2), We will also prioritize by calculated properties that correlate with cell permeability. The top 200 compounds from this virtual screening campaign will be purchased, and in vitro activity will be evaluated according to a tiered progression scheme. Compounds with good potency in an enzyme assay (Ki < 10µM) will be advanced to PAMPA for assessing permeability, and then MT1-2 receptor assays for selectivity. Proof-of-concept efficacy studies in zebrafish larvae will provide an efficient and validated way to evaluate SNAT inhibition in a living system. In Aim 2, we will use our model to design new scaffolds based on one of the few existing drug-like SNAT inhibitors. Also, we expand upon an innovative but insufficiently studied strategy for forming inhibitors at SNAT’s active site via its alkyltransferase activity. Powerful kinetic mechanism methods and co-crystal structures of selected inhibitors will be employed to develop a more complete model of SNAT inhibition, which would be highly impactful for studying and treating CR disorders.
